DNA-Based Optical Quantification of Ion Transport across Giant Vesicles.

Accurate measurements of ion permeability through cellular membranes remains challenging due to the lack of suitable ion-selective probes. Here we use giant unilamellar vesicles (GUVs) as membrane models for the direct visualization of mass translocation at the single-vesicle level. Ion transport is indicated with a fluorescently adjustable DNA-based sensor that accurately detects sub-millimolar variations in K+ concentration. In combination with microfluidics, we employed our DNA-based K+ sensor for extraction of the permeation coefficient of potassium ions. We measured K+ permeability coefficients at least 1 order of magnitude larger than previously reported values from bulk experiments and show that permeation rates across the lipid bilayer increase in the presence of octanol. In addition, an analysis of the K+ flux in different concentration gradients allows us to estimate the complementary H+ flux that dissipates the charge imbalance across the GUV membrane. Subsequently, we show that our sensor can quantify the K+ transport across prototypical cation-selective ion channels, gramicidin A and OmpF, revealing their relative H+/K+ selectivity. Our results show that gramicidin A is much more selective to protons than OmpF with a H+/K+ permeability ratio of ∼104.

Identification of GmGPATs and their effect on glycerolipid biosynthesis through seed-specific expression in soybean.

BACKGROUND: Genetic improvement of soybean oil content depends on in-depth study of the glycerolipid biosynthesis pathway. The first acylation reaction catalysed by glycerol-3-phosphate acyltransferase (GPAT) is the rate-limiting step of triacylglycerol biosynthesis. However, the genes encoding GPATs in soybean remain unknown. METHODS: We used a novel yeast genetic complementation system and seed-specific heterologous expression to identify GmGPAT activity and molecular function in glycerolipid biosynthesis. RESULTS: Sixteen GmGPAT genes were cloned by reverse transcription-PCR for screening in yeast genetic complementation. The results showed that GmGPAT9-2 could restore the conditional lethal double knockout mutant strain ZAFU1, and GmGPAT1-1 exhibited low acyltransferase activity in serial dilution assays. In addition, the spatiotemporal expression pattern of GmGPAT9-2 exhibited tissue specificity in leaves, flowers and seeds at different developmental stages. Furthermore, both the proportion of arachidic acid and erucic acid were significantly elevated in Arabidopsis thaliana transgenic lines containing the seed-specific GmGPAT9-2 compared wild type, but the oil content was not affected. CONCLUSION: Together, our results provide reference data for future engineering of triacylglycerol biosynthesis and fatty acid composition improvement through GPATs in soybean.

Tailoring the Binding Properties of Phosphazane Anion Receptors and Transporters.

The binding and sensing of anions is an important cross-disciplinary field, which impacts broad areas such as biology, supramolecular chemistry and catalysis. To date, however, this area has been dominated by organic architectures which function as H-bonding, anion receptor molecules. Inorganic anion receptors have largely been based on Lewis acidic metals, with very few examples of H-bonding counterparts of organic systems having been systematically studied. This paper develops strategies for enhancing the anion binding properties of phosphazanes of the type [(RNH)(E)P(µ-N tBu)]2 (E = O, S, Se) which are bench-stable, H-bond receptors that can be regarded as inorganic analogues of squaramides (a key class of organic anion receptor). The distinct advantages of these inorganic receptors over organic counterparts is the ease by which their functionality and electronic character can be altered (by means of the R group, chalcogenide, or metal present). Se substitution at the P centers, the presence of electron-withdrawing R groups, and metal coordination to the soft donor centers can be used to modulate and enhance anion binding. The water stability and superior anion binding properties of the seleno-phosph(V)azanes give them applications as synthetic anion transporters through phospholipid layers.